Transcriptional profiling distinguishes inner and outer annulus fibrosus from nucleus pulposus in the bovine intervertebral disc

• Published in: European spine journal Vol. 26; no. 8; pp. 2053 - 2062
• Main Authors: van den Akker, Guus G. H., Koenders, Marije I., van de Loo, Fons A. J., van Lent, Peter L. E. M., Blaney Davidson, Esmeralda, van der Kraan, Peter M.
• Format: Journal Article
• Language: English
• Published: Berlin/Heidelberg Springer Berlin Heidelberg 01-08-2017; Springer Nature B.V
• Subjects: Animals; Annulus Fibrosus - metabolism; Cartilage (articular); Cattle; Gene expression; Gene Expression Profiling; Genetic Markers; Intervertebral discs; Medicine; Medicine & Public Health; Neurosurgery; Nuclei; Nucleus pulposus; Nucleus Pulposus - metabolism; Original Article; Phenotype; Surgical Orthopedics; Transcription; Transcriptome; Cell biology; Annulus fibrosus; Nucleus pulposus; Intervertebral disc
• ISSN: 0940-6719; 1432-0932
• DOI: 10.1007/s00586-017-5150-3

Background
Cells in the intervertebral disc have unique phenotypes and marker genes that separate the nucleus pulposus (NP), annulus fibrosus (AF) and articular cartilage (AC) have been identified. Recently, it was shown that phenotypic marker genes exhibit variable expression in humans. In this study, the bovine tail was used to determine the ability of marker genes to distinguish the outer and inner AF from NP tissue and isolated cells.
Methods
Bovine tail intervertebral discs from 13 donors were dissected and correct isolation of tissue was confirmed. mRNA was isolated directly from tissue or passage 0 monolayer cells and used for gene expression measurements (qPCR). Conventional marker genes (
bAcan, bCol1a1, bCol2a1
) and novel marker genes (
bAdamts17, bBrachyury/T, bCD24, bCol5a1, bCol12a1, bFoxf1, bKrt19, bPax1, bSfrp2
) were evaluated.
Results
As expected
bAcan
,
bCol2a1
and
bCol1a1
distinguished outer AF from NP tissue, while inner AF and NP could not be discriminated. The NP markers
bT, bCd24
and
bKrt19
were significantly higher expressed in NP than inner and outer AF tissue.
bFoxF1
and
bPax1
only distinguished IVD tissues from AC. The AF markers
bAdamts17, bCol5a1, bCol12a1
and
bSfrp2
were higher expressed in the outer AF compared with inner AF and NP tissue. Monolayer culturing strongly decreased
bAcan, bCol2a1, bCD24
and
bCol5a1
expression, while
bCol1a1, bT, bKrt19
and
bSfrp2
were not affected.
Conclusion
The IVD phenotypic marker genes
bT, bKrt19, bSfrp2
and
bCol12a1
convincingly distinguished NP from outer AF in situ and in vitro.